During late pregnancy in humans and other mammalian species, the consistency of the cervix softens, thus enabling cervical dilatation and rapid delivery of the fetus(es). In cases of difficult spontaneous delivery or induced delivery at term, a hard inelastic cervix may be a source of complications. The hormone relaxin is an effective cervical softening agent with potential for clinical application in women and domestic animals. Informed decisions concerning the use of relaxin as a cervical softening agent will largely depend upon basic studies of its effects in species other than the human. The principal long-range objective of this application is to better understand at the cellular and molecular levels the mechanism(s) whereby relaxin, in concert with estrogen, promotes growth and softening of the cervix in pregnant rats. To determine the influence of estrogen and relaxin on (1) the composition and ultrastructure, as well as (2) the proliferation of cells within the cervix, rats will be ovariectomized on day 9 of pregnancy and thereafter receive hormone replacement therapy with physiological levels of progesterone, progesterone and estrogen, or progesterone, estrogen, and relaxin. Cervices will be collected late in pregnancy (day 22) and the effects of estrogen and relaxin on the cervical cells will be determined by light and electron microscopy. Immunohistochemistry will be used to (3) identify the target cells for relaxin in the cervix. Ovariectomized hormone-treated pregnant rats will also be used for studies aimed at determining the influence of estrogen and relaxin on enzymes (prostaglandin synthase, collagenase, stromelysin) synthesized by rat cervices. The quantities of enzymes within cervical tissues will be determined by Western blot analysis and the cellular localization of the enzymes will be done by immunocytochemical analysis. The quantities of mRNA for the cervical enzymes will be determined by ribonuclease protection assay and the localization of the mRNA for enzymes will be done by in situ hybridization. Specific Aim 5 of this application is to determine if endogenous relaxin has physiological effects on the vagina in pregnant rats. The consequences of neutralization of relaxin with a monoclonal antibody for rat relaxin on the growth, tensile properties, and histological characteristics of the vagina will be determined.